Since the first mutations of the neuronal sodium channel SCN1A were identified 5 years ago, more than 150 mutations have been described in patients with epilepsy. Many are sporadic mutations and cause loss of function, which demonstrates haploinsufficiency of SCN1A. Mutations resulting in persistent sodium current are also common. Coding variants of SCN2A, SCN8A, and SCN9A have also been identified in patients with seizures, ataxia, and sensitivity to pain, respectively. The rapid pace of discoveries suggests that sodium channel mutations are significant factors in the etiology of neurological disease and may contribute to psychiatric disorders as well.

Epilepsy is a common neurological disorder affecting approximately 1% of the population. Mutations in voltage-gated sodium channels are responsible for several monogenic epilepsy syndromes. More than 800 mutations in the voltage-gated sodium channel SCN1A have been reported in patients with generalized epilepsy with febrile seizures plus and Dravet syndrome. Heterozygous loss-of-function mutations in SCN1A result in Dravet syndrome, a severe infant-onset epileptic encephalopathy characterized by intractable seizures, developmental delays and increased mortality. A common feature of monogenic epilepsies is variable expressivity among individuals with the same mutation, suggesting that genetic modifiers may influence clinical severity. Mice with heterozygous deletion of Scn1a (Scn1a+/−) model a number of Dravet syndrome features, including spontaneous seizures and premature lethality. Phenotype severity in Scn1a+/− mice is strongly dependent on strain background. On the 129S6/SvEvTac strain Scn1a+/− mice exhibit no overt phenotype, while on the (C57BL/6J × 129S6/SvEvTac)F1 strain Scn1a+/− mice exhibit spontaneous seizures and early lethality. To systematically identify loci that influence premature lethality in Scn1a+/− mice, we performed genome scans on reciprocal backcrosses. QTL mapping revealed modifier loci on mouse chromosomes 5, 7, 8 and 11. RNA-seq analysis of strain-dependent gene expression, regulation and coding sequence variation provided a list of potential functional candidate genes at each locus. Identification of modifier genes that influence survival in Scn1a+/− mice will improve our understanding of the pathophysiology of Dravet syndrome and may suggest novel therapeutic strategies for improved treatment of human patients.

Heterozygous loss-of-function SCN1A mutations cause Dravet syndrome, an epileptic encephalopathy of infancy that exhibits variable clinical severity. We utilized a heterozygous Scn1a knockout (Scn1a+/−) mouse model of Dravet syndrome to investigate the basis for phenotype variability. These animals exhibit strain-dependent seizure severity and survival. Scn1a+/− mice on strain 129S6/SvEvTac (129.Scn1a+/−) have no overt phenotype and normal survival compared with Scn1a+/− mice bred to C57BL/6J (F1.Scn1a+/−) that have severe epilepsy and premature lethality. We tested the hypothesis that strain differences in sodium current (INa) density in hippocampal neurons contribute to these divergent phenotypes. Whole-cell voltage-clamp recording was performed on acutely-dissociated hippocampal neurons from postnatal day 21–24 (P21–24) 129.Scn1a+/− or F1.Scn1a+/− mice and wild-type littermates. INa density was lower in GABAergic interneurons from F1.Scn1a+/− mice compared to wild-type littermates, while on the 129 strain there was no difference in GABAergic interneuron INa between 129.Scn1a+/− mice and wild-type littermate controls. By contrast, INa density was elevated in pyramidal neurons from both 129.Scn1a+/− and F1.Scn1a+/− mice, and was correlated with more frequent spontaneous action potential firing in these neurons, as well as more sustained firing in F1.Scn1a+/− neurons. We also observed age-dependent differences in pyramidal neuron INa density between wild-type and Scn1a+/− animals. We conclude that preserved INa density in GABAergic interneurons contributes to the milder phenotype of 129.Scn1a+/− mice. Furthermore, elevated INa density in excitatory pyramidal neurons at P21–24 correlates with age-dependent onset of lethality in F1.Scn1a+/− mice. Our findings illustrate differences in hippocampal neurons that may underlie strain- and age-dependent phenotype severity in a Dravet syndrome mouse model, and emphasize a contribution of pyramidal neuron excitability.

ObjectiveEpilepsy is a common neurological disorder that affects 1% of the population. Approximately, 30% of individuals with epilepsy are refractory to treatment, highlighting the need for novel therapies. Conventional anticonvulsant screening relies predominantly on induced seizure models. However, these models may not be etiologically relevant for genetic epilepsies. Mutations in SCN1A are a common cause of Dravet Syndrome, a severe epileptic encephalopathy. Dravet syndrome typically begins in infancy with seizures provoked by fever and then progresses to include afebrile pleomorphic seizure types. Affected children respond poorly to available anticonvulsants. Scn1a +/− heterozygous knockout mice recapitulate features of Dravet syndrome and provide a potential screening platform to investigate novel therapeutics. In this study, we conducted a screening of conventional anticonvulsants in Scn1a +/− mice to establish assays that most closely correlate with human response data.MethodsOn the basis of clinical response data from a large, single center, retrospective survey of Dravet syndrome case records, we selected nine drugs for screening in Scn1a +/− mice to determine which phenotypic measures correlate best with human therapeutic response. We evaluated several screening paradigms and incorporated pharmacokinetic monitoring to establish drug exposure levels.Results Scn1a +/− mice exhibited responses to anticonvulsant treatment similar to those observed clinically. Sodium channel blockers were not effective or exacerbated seizures in Scn1a +/− mice. Overall, clobazam was the most effective anticonvulsant in Scn1a +/− mice, consistent with its effect in Dravet syndrome.InterpretationGenetic models of spontaneous epilepsy provide alternative screening platforms and may augment the AED development process. In this study, we established an effective screening platform that pharmacologically validated Scn1a +/− mice for preclinical screening of potential Dravet syndrome therapeutics.

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Autism is a psychiatric disorder with estimated heritability of 90%. One-third of autistic individuals experience seizures. A susceptibility locus for autism was mapped near a cluster of voltage-gated sodium channel genes on chromosome 2. Mutations in two of these genes, SCN1A and SCN2A, result in the seizure disorder GEFS þ . To evaluate these sodium channel genes as candidates for the autism susceptibility locus, we screened for variation in coding exons and splice sites in 117 multiplex autism families. A total of 27 kb of coding sequence and 3 kb of intron sequence were screened. Only six families carried variants with potential effects on sodium channel function. Five coding variants and one lariat branchpoint mutation were each observed in a single family, but were not present in controls. The variant R1902C in SCN2A is located in the calmodulin binding site and was found to reduce binding affinity for calcium-bound calmodulin. R542Q in SCN1A was observed in one autism family and had previously been identified in a patient with juvenile myoclonic epilepsy. The effect of the lariat branchpoint mutation was tested in cultured lymphoblasts. Additional population studies and functional tests will be required to evaluate pathogenicity of the coding and lariat site variants. SNP density was 1/kb in the genomic sequence screened. We report 38 sodium channel SNPs that will be useful in future association and linkage studies.

Background Numerous studies have demonstrated increased load of de novo copy number variants (CNVs) or single nucleotide variants (SNVs) in individuals with neurodevelopmental disorders, including epileptic encephalopathies, intellectual disability and autism. Methods We searched for de novo mutations in a family quartet with a sporadic case of epileptic encephalopathy with no known etiology to determine the underlying cause using high coverage whole exome sequencing (WES) and lower coverage whole genome sequencing (WGS). Mutations in additional patients were identified by WES. The effect of mutations on protein function was assessed in a heterologous expression system. Results We identified a de novo missense mutation in KCNB1 that encodes the KV2.1 voltage-gated potassium channel. Functional studies demonstrated a deleterious effect of the mutation on KV2.1 function leading to a loss of ion selectivity and gain of a depolarizing inward cation conductance. Subsequently, we identified two additional patients with epileptic encephalopathy and de novo KCNB1 missense mutations that cause a similar pattern of KV2.1 dysfunction. Interpretation Our genetic and functional evidence demonstrate that KCNB1 mutation can result in early onset epileptic encephalopathy. This expands the locus heterogeneity associated with epileptic encephalopathies and suggests that clinical WES may be useful for diagnosis of epileptic encephalopathies of unknown etiology.

Dravet syndrome is a severe, childhood-onset epilepsy largely due to heterozygous loss-of-function mutation of the gene , which encodes the type 1 neuronal voltage-gated sodium (Na) channel α subunit Nav1.1. Prior studies in mouse models of Dravet syndrome ( mice) indicate that, in cerebral cortex, Nav1.1 is predominantly expressed in GABAergic interneurons, in particular in parvalbumin-positive fast-spiking basket cell interneurons (PVINs). This has led to a model of Dravet syndrome pathogenesis in which Nav1.1 mutation leads to preferential dysfunction of interneurons, decreased synaptic inhibition, hyperexcitability, and epilepsy. However, such studies have been implemented at early developmental time points. Here, we performed electrophysiological recordings in acute brain slices prepared from male and female mice as well as age-matched wild-type littermate controls and found that, later in development, the excitability of PVINs had normalized. Analysis of action potential waveforms indirectly suggests a reorganization of axonal Na channels in PVINs from mice, a finding supported by immunohistochemical data showing elongation of the axon initial segment. Our results imply that transient impairment of action potential generation by PVINs may contribute to the initial appearance of epilepsy, but is not the mechanism of ongoing, chronic epilepsy in Dravet syndrome. Dravet syndrome is characterized by normal early development, temperature-sensitive seizures in infancy, progression to treatment-resistant epilepsy, developmental delay, autism, and sudden unexplained death due to mutation in encoding the Na+ channel subunit Nav1.1. Prior work has revealed a preferential impact of Nav1.1 loss on the function of GABAergic inhibitory interneurons. However, such data derive exclusively from recordings of neurons in young mice. Here, we show that impaired action potential generation observed in parvalbumin-positive fast-spiking interneurons (PVINs) in+/- mice during early development has normalized by postnatal day 35. This work suggests that a transient impairment of PVINs contributes to epilepsy onset, but is not the mechanism of ongoing, chronic epilepsy in Dravet syndrome.